The present invention relates to the novel use of a new, low phosphorus, low-pigmented (xanthophyll), highly digestible white protein gluten meal to be used as an ingredient in feeding operations, especially aquaculture. It is derived from the wet milling of an identity preserved, white corn hybrid with distinct characteristics. The development of this novel product involves several selection criteria across a variety of disciplines involving plant breeding, industrial processing, feeding operations, and environmental issues.
The publications and other materials used herein to illuminate the background of the invention or provide additional details respecting the practice, are incorporated by reference, and for convenience are respectively grouped in the appended Bibliography.
The goal of plant breeding is to develop new, unique, and superior corn inbred lines and hybrids. Typically, the development of a hybrid corn variety involves three steps: 1) the selection of plants from various germplasm pools for initial breeding crosses; 2) the selfing of the selected plants from the breeding crosses for several generations to produce a series of inbred lines, which, although different from each other, breed true and are highly uniform; and, 3) crossing the selected inbred lines with unrelated inbred lines to produce the hybrid progeny (F1). The breeder can theoretically generate billions of different genetic combinations via crossing, selfing, and mutations.
One of the most important tasks corn breeders have is to evaluate newly developed experimental materials. The difficulty in this task is to separate genetic and environmental effects. The usual procedure is to evaluate the material in performance trials conducted over two or three years at a minimum of six locations. More locations are preferred, but the resources available determine the number. Testing sites should be located in areas where the newly developed material is likely to be marketed.
Most plant breeders use a screening type of trial for eliminating genotypes that are obviously poor. Usually, large numbers of many genotypes are observed at a few locations. Sometimes, inoculations with prevalent leaf diseases and stalk rot pathogens are included. Those hybrids that survive rigorous testing are usually grown in larger strip tests for evaluation by farmers. If reaction is favorable, the selected hybrids are put into pilot seed production, and also entered in state variety trials before being placed into large-scale production.
Corn kernels can be altered by genetic means to give modifications in starch, protein, oil, pericarp thickness, kernel hardness, embryo size, kernel size, and color. This generates another set of testing parameters for products that are designed to be used by corn processors for specific value-added products or co-products.
Starch from normal dent or flint corn is composed of 73% amylopectin (the starch fraction with branched molecules) and 27% amylose (the fraction with linear molecules). Waxy corn (having the wx gene) was first found in China, but waxy mutations have also been found in American dent strains. Starch from this mutation is 100% amylopectin.
The endosperm mutant amylose-extender (ae) was found by R. P. Bear in 1950 (Vineyard et al., 1958). It increased the amylose fraction of the starch to 50% and above. The kernel of this corn is characterized by a tarnished, translucent, and partially full appearance. The ae gene, plus modifiers, gives a range in amylose content of 50-80%, but the amylose content can be stabilized at various intermediate levels.
Several endosperm mutants that alter the balance of amino acids have been identified. The most important of these is opaque-2. Mertz et al. (1964) reported that opaque-2 reduced zein in the endosperm and increased lysine. Other mutant genes with similar effects are floury-2 and opaque-7.
Kernels with the opaque-2 gene are characterized by a soft, chalky, nontransparent appearance, with very little hard vitreous or horney endosperm. This type of kernel is more prone to damage by kernel rots, insects, rodents, and harvesting machinery.
Another source of increased lysine and protein in single cross hybrids was discovered by Strissel et al. (U.S. Pat. No. 5,082,993). The increased nutritional levels were inherited in a dominant manner verses the recessive nature of the opaque-2 gene, and it was not a mutation. The inbred that was patented was derived from an exotic germplasm source, and not only increased protein levels, but the protein was also more digestible.
The mature corn kernel has four easily separable parts: tip cap, pericarp, endosperm, and germ. The major component of corn is starch, of which 98% is in the endosperm (Earle et al., 1946). On the whole kernel basis, starch content is 72-73%. The endosperm also contains 74% of the kernel protein, of which the majority is insoluble storage proteins.
The germ is the major depository of lipids, which amount to 83% of the total kernel lipids. The greater parts of the germ lipids are triacylglycerides, which, on extraction, give the well-known corn oil of commerce. The germ, being potentially metabolically active tissue, contains 70% of the kernel sugar and 26% of the kernel protein. Most of the germ proteins are albumins or globulins and probably are components of the enzymatic apparatus of the cells.
The corn germ is also rich in mineral elements that are essential for early growth of the embryo. The embryo contains 78% of the kernel minerals of which inorganic phosphorus is the most abundant. It is largely present as the potassium-magnesium salt of phytic acid-the hexaphosphate ester of inositol. Phytin is an important storage form of phosphorus (Hamilton et al., 1951; O""Dell et al., 1972), which is liberated by phytase enzymes to initiate embryo development. More than 80% of the phosphorus in corn is in the form of phytate. The corn germ contains nearly 90% of the phytate present in whole corn.
One of the problems associated with the use of grain and oilseed products in feed for monogastric animals, e.g. pigs, chickens, and fish, is the presence of phytate. Phytate phosphorus is nutritionally unavailable, and is excreted in the feces. It is then suspected of contributing to nutrient enrichment of several ecosystems when manure from confined animal rearing operations leaches into the ground and from there into lakes, streams, and bays.
In catfish rearing, excretion of phytate, which is degraded by microorganisms, thus releasing the bound phosphorus, contributes to algae growth in ponds. In trout farming, reducing the phosphorus levels of feeds over the past few years has greatly lowered the amount of soluble phosphorus excreted via urine, but little change has occurred in the amount of insoluble phosphorus excreted in the feces.
In trout feeds containing 1.6% total phosphorus, phytate phosphorus makes up 0.22% of total phosphorus. Feed manufacturers are reducing the amount of fish meal and replacing it with soy, wheat, and corn-based protein concentrates. These concentrates are lower in total phosphorus, but also lower in available phosphorus because of phytate. In a trout feed in which 2/3 of the fish meal is replaced with plant protein sources, phytate phosphorus could make up half or more of the total phosphorus in the feed, increasing fecal phosphorus loss.
Phytate has other possible effects on feed constituents mainly associated with its ability to interact directly and indirectly with certain minerals especially calcium, magnesium, zinc, and iron to reduce their availability to animals (Underwood, 1962; Momcilovic and Shahl, 1976). For example, calcium-bound phytate increases chelation with trace minerals, especially zinc, to form co-precipitates that make the zinc unavailable to the animal or fish.
Richardson et al. (1985) showed that zinc availability was greatly reduced to juvenile chinook salmon when sodium phytate was added to their feed, and that the fish developed cataracts as a result. In channel catfish, just 1.1% supplemental phytate in the feed increased the amount of zinc in the diet necessary to prevent deficiency signs by 10xc3x97 over the amount needed in a semi-purified diet (Gatlin and Wilson, 1984).
Phosphorus content of common fish feed ingredients is highly variable. Some practical ingredients contain limited amounts of P (e.g. 0.3% P in blood meal) while other contain very significant levels (4-5% P in meat and bone meal) (NRC, 1993). Phosphorus contained in organic compounds, such as phospholipids and nucleic acids, are apparently highly digestible for fish. Phosphorus contained in phytate (inositol hexaphosphate), also an organic compound, is not digestible to fish since they lack the necessary enzyme (phytase). The digestibility of mineral forms of P, such as dicalcium phosphate, monosodium phosphate and rock phosphate, varies with degree of solubility of the compound(s) and is, consequently, highly variable (Lall, 1991). The digestibility of P contained in bone (apatite) is variable between fish species and depends mostly on stomach pH of the animal (Lall, 1991). For rainbow trout, a fish with a true (acid) stomach, apparent digestibility coefficient (ADC) of bone P appears to be between 40 and 60%. ADC of bone P appears to be much lower for stomachless fish, such as carp (Lall, 1991). Other factors, such as particle size, feed processing technique, and enzyme treatment, are also known to affect ADC of P (Lall, 1991).
Numerous studies have shown that dietary incorporation of microbial phytase improved the ADC of P of fish fed diets containing phytic acid (Rodehutscord and Pfeffer, 1995; Oliva-Teles et al., 1998; Vielma et al., 1998; Forster et al., 1999). The activity of this enzyme is affected by environmental temperature and its activity may be very limited at low water temperatures (Forester et al., 1999). Moreover, the enzyme is sensitive to heat and may be destroyed during pelleting and extrusion under standard commercial conditions.
Both digestibility and quantity will determine the fate of P fed to fish. The undigested fraction of the P of the diet is excreted in the feces by fish. The fraction of P digested by the animal is absorbed where it is deposited in the body of the fish (bones, scales, flesh, etc.) in the growth processes. A number of experimental evidences suggest that there is a requirement to maximize growth and maximize phosphorus deposition and bone mineralization. Phosphorus requirement of rainbow trout for maximum growth was 0.37% digestible P and 0.53% for maximum phosphorus deposition (Rodehutscord, 1996).
Fish receiving only the digestible P amount required to meet requirement for growth excrete only minute amounts of non-fecal P indicating that digestible P intake of the fish is directed almost completely toward deposition (Rodehutscord, 1996; Vielma and Lall, 1998; Bureau and Cho, 1999). There is evidence that efficiency of P utilization tends to decrease as digestible P level increase from the level required for maximum growth to the level required for maximum P deposition (Rodehutscord, 1996; Rodehutscord et al., 2000b). Interpretation of available data suggest that, while feeding a diet with digestible P at the level required to maximize growth results in minimal non-fecal P excretion, feeding with a diet with a digestible P level required that maximum P deposition results in significant non-fecal excretion.
Fish meal has been the main protein source used in feeds for farmed carnivorous fish since fish farming began. Fish meal is produced from stocks of fish not harvested for human consumption, and annual global production is 6.5 million metric tons (mmt) per year (Hardy and Green, 1999). Peru and Chile have traditionally accounted for approximately 35% of annual global production, utilizing anchovies and horse mackerel. Periodically, production from Peru and Chile declines by over 80%, in El Nino years when ocean temperatures and currents cause the fish stocks to move offshore, out of reach of the fishery. In years when this occurs, world production of fish meal declines, generally to 4-4.5 mmt, and the cost of fish meal increases to over $600/mt, double its lowest value. If other fish meal-producing countries are experiencing declines in their fisheries during El Nino years, the supply/demand situation becomes even worse.
When global fish meal production declines, feed producers turn to alternative proteins, such as rendered products and protein concentrates from grains and oilseeds. Among rendered products, poultry by-product meal, meat and bone meal, blood meal, and feather meal are utilized in animal and fish feeds. Soybean meal (de-hulled) is the main protein source from oilseeds that is used in animal feeds. Corn gluten meal is the main protein source from grains. Other substitutes include canola meal, soy protein concentrate, and wheat gluten, although price limits use of soy protein concentrate and wheat gluten (Hardy, 1999).
Feeds for aquatic animals now consume over 25% of global fish meal production each year, up from 10-12% eight years ago (Hardy, 1999). Poultry feeds use 50% of global production, swine 15%, and the remainder is used in pet feeds, rumen by-pass proteins, and minor species feeds. Continued growth of aquaculture is expected to increase the proportion of global fish meal used in feeds for fish and shrimp, according to projections. However, most observers assume the percentage of fish meal used in fish feeds will decrease. Underlying this assumption is the second assumption that alternative protein sources will substitute for fish meal. At present, feeds for three species groups in aquaculture account for 70% of fish meal use in aquaculture feeds: salmon, shrimp, and trout.
Yellow corn gluten meal has been evaluated as a fish meal substitute for fish with moderate success. Weede (1997) found that corn gluten could substitute for 25% of the fish meal in feeds for rainbow trout, but that 50% substitution or higher resulted in reduced feed intake and growth. Skonberg et al. (1998) reported that the use of over 15% corn gluten meal in trout feeds caused a yellowing of the flesh, and most trout feed manufacturers limit corn gluten meal to 5% in feeds or avoid it altogether. Corn gluten protein is reported to be relatively well digested by rainbow trout and Pacific salmon, but phosphorus availability is low (Sugiura et al., 1998), likely the result of phytate. Thus, what limits the use of corn gluten meal in feeds for salmonoids and shrimp is the presence of xanthophyll pigments and phytate. If these problems could be overcome, the potential market, at 25% use in the feed, is approximately 900,000 mt for salmon, shrimp, and trout. At least double this potential market exists in other species, notably European sea bass, gilthead sea bream, yellowtail, and various freshwater catfish species in Asia.
Yellow corn gluten meal is derived from the wet-milling process of corn. The primary products from the wet-milling process are food and industrial starches and sweeteners. Co-products include corn oil and the feed products corn gluten feed (CGF), corn gluten meal (CGM), corn germ meal, and condensed fermented corn extractives (steep liquor). The germ is solvent-extracted to recover oil, and the extracted germ meal is used in feed products. The gluten is separated from starch by centrifuges, giving a stream containing 69-72% (dry substance basis) total protein, which is dried to become 60% protein CGM. It is highly digestible, contains metabolizable energy (ME) of 4,131 kcal/kg of dry matter for the chick, and is rich source of available carotenes (49-73 mg/kg) and xanthophylls (244-550 mg/kg, dry substance basis). Its crude protein is highly digestible, a good source of methionine and cystine, but very low in lysine and tryptophan. The average phosphorus content of the yellow gluten meal is 0.78% (dry substance basis) (Corn: Chemistry and Technology, 1987). The solubles removed from the corn during steeping are concentrated by evaporation and are called steep liquor. The corn bran (fiber), corn germ meal, and the steep liquor are combined to form CGF with a typical protein content of 18-21%.
Normal yellow dent corn is the predominant corn processed. Its starch has 27% amylose, a linear glucose polymer, and 73% amylopectin, a branch-chain glucose polymer.
Flint corn, a variety of normal corn grown in South America, is processed successfully even though it does not soften much, even with 50-60 hours of steeping. Its starch yield is slightly less than regular corn, but its quality is good and the amylose-amylopectin ratio is the same as that of normal dent corn. No flint corn is processed in the United States.
Waxy corn, a genetic mutant, contains starch composed entirely of amylopectin. It can be processed in the same way as regular corn, with minor adjustments. The pasting temperature is lower, so the process must be cooler by about 3xe2x96xa1 C. Separation of the starch and gluten is easier, but the starch yield is only 90% of that of regular corn. About 1.5% of the corn processed by wet milling in the United States is the waxy type.
Another genetic mutant variety that is processed commercially is high-amylose corn, with starch that is 60-70% amylose. Like waxy corn, the farmer must grow it in fields isolated from other varieties to prevent cross-pollination and then handle it separately after harvest to avoid contamination. Unlike the lower temperature for waxy corn, the processing temperature should be 3xe2x96xa1 C. higher than that for regular corn. More steep time is required, starch-gluten separation and starch filtration are more difficult, and the starch yield is only 80-90% of that of regular corn. Only about 0.2% of the corn wet milled in the United States is the high-amylose variety.
White corn hybrids are mostly dry milled or used for other food products, and usually bred for hard endosperm, making them unsuitable for industrial wet milling purposes. A small quantity of white corn has traditionally been wet milled to produce specialty products with very bright whiteness. To be economically feasible, the value of the resultant specialty starch not only has to overcome the production premium but must also compensate for the decreased value of the gluten meal. A premium is paid in the U.S. market for gluten meal because the yellow carotenoids are desired by the poultry industry. The value of the pigmentation is considered to be $0.03-$0.07 per lb. of gluten meal or $0.08-$0.19 per bushel.
Traditionally, the dry and wet milling industries have selected varieties based on basic items such as kernel hardness, color, amount of stress cracks, thins, test weight, grit to germ ratio, density, and pericarp removal. All approved hybrids are typically collected and processed in a random manner.
The invention described herein relates to the novel use of a white protein gluten meal derived from the wet milling of one or more identity-preserved white corn hybrids with specific traits.
The present invention includes the use of a white protein gluten meal in the aquaculture feed industry with possible applications for additional livestock operations. The use of this new ingredient has implications on the quality of the meat, nutrient deposition in the meat, and potential positive environmental effects.
The present invention further includes selecting hybrid varieties of white corn that have specific grain components, for example, protein characteristics of the endosperm, starch characteristic, xanthophylls, phosphorus, and wet milling traits related to the separation and development of white protein gluten meal.
Thus, in a first aspect, the present invention is directed to white protein gluten meal having a phosphorus content in the range of about 0.30% to about 0.60%. The white protein gluten meal may also have a xanthophyll content of less than 80 mg/Kg in the range of about 2.5 mg/Kg to about 80.0 mg/Kg. The white protein gluten meal may further have a beta-carotene content of less than 3.0 mg/Kg with a range of about 0.03 to about 3.0 mg/Kg. Apparent digestibility coefficients (ADC) for the protein of the present invention are in the range of about 80.0% to about 96.0%. Finally, the white protein gluten meal has a crude protein content greater than 70% on a dry substance basis (DSB) in the range of about 70.0% to about 85.0%.
In a second aspect, the present invention is directed to a method for producing a white corn gluten meal comprising: a) selecting white corn grain having a total grain phosphorus content of less than 0.28%; and b) wet milling said grain to produce a white protein gluten meal. The white corn grain may also have a xanthophyll content of less than 5.3 mg/Kg with a range of between about 2.0 mg/Kg to about 5.3 mg/Kg. The white corn may further have a beta-carotene content of less than 0.10 ug/10 g with a range of between about 0.02 ug/100 g to about 0.10 ug/100 g. Finally, the white corn is selected such that the crude protein content after wet milling is greater than 70% (DSB) in the range of between about 70% to about 85%.
In a third aspect, the present invention is directed to an animal feed containing 1% to 60% white protein gluten meal. The animal feed preferably contains 12% to 24% white protein gluten meal. The animal feed more preferably contains 24% to 50% white protein gluten meal. The animal feed is particularly useful in applications that need a highly digestible protein source with increased availability of phosphorus, and the absence of xanthophyll pigments. One such application is aquaculture.
In a fourth aspect, the present invention is directed to a method for animal husbandry which comprises feeding animal feed containing 1% to 60% white protein gluten meal to an animal. The animal feed preferably contains 12% to 24% white protein gluten meal. The animal feed more preferably contains 24% to 50% white protein gluten meal. In a preferred embodiment, the animal husbandry is aquaculture.
In a fifth aspect, the present invention is directed to a method for reducing loss of phosphorus in an animal feeding operation comprising feeding animal feed containing 1% to 60% white protein gluten meal to an animal. The animal feed preferably contains 12% to 24% white protein gluten meal. The animal feed more preferably contains 24% to 50% white protein gluten meal. In a preferred embodiment, the animal feeding operation is aquaculture.
In a sixth aspect, the present invention is directed to a method for increasing phosphorus retention in animal flesh in an animal feeding operation comprising feeding animal feed containing 1% to 60% white protein gluten meal to an animal. The animal feed preferably contains 12% to 24% white protein gluten meal. The animal feed more preferably contains 24% to 50% white protein gluten meal. In a preferred embodiment, the animal feeding operation is aquaculture.
The invention includes the use of a new ingredient, white protein gluten meal, in aquaculture feeding operations to replace fish meal as an alternative protein source that lowers total phosphorus input and increases phosphorus retention in the flesh. In addition, this product is nearly devoid of xanthophyll pigments, and generates white fish fillets verses the yellow pigmented fillets wherever yellow gluten is used at levels greater than 6%.
The following definitions are used herein.
Beta-carotene content: The total amount of beta-carotene. Beta-carotene has a 40 carbon base, and is the dominant provitamin A carotenoid in corn.
Corn Gluten Feed (CGF): A feed ingredient with a medium protein level and is palatable to all classes of livestock and poultry. CGF does not contain any gluten. It is comprised of the fiber fraction (bran), steep liquor, and, where available, germ meal. It commonly contains a minimum of 21% crude protein and approximately 15% starch.
Corn Gluten Meal (CGM): A feed ingredient that is the dehydrated protein stream resulting from starch separation in the endosperm fraction of the grain. It has a high nutrient density and usually is sold containing a minimum of 60% total protein. It is highly digestible, and is a rich source of carotenes (49-73 mg/kg) and xanthophylls (244-550 mg/kg).
Crude Protein Content: The total nitrogenous material in the plant substance. Protein=Nxc3x976.25.
Dry Substance Basis (DSB): Measurements are based on a zero moisture content basis.
Feed Efficiency (FE): As used herein, the feed efficiency in a feeding study is determined by dividing the wet weight gain by the dry feed intake. Diets with the highest number would have the best and highest feed efficiency.
Identity Preserved White Corn Hybrid with Specific Traits: White corn hybrid grain produced by a system where corn growers purchase a specific white corn hybrid, grow it in isolation to prevent contamination from yellow or a different white corn, dry the grain with a lower temperature to keep the grain temperature less than 140xe2x96xa1 F. and then store the grain in separate bins to maintain purity.
Phosphorus content: The total amount of the mineral element phosphorus. It is largely present as the potassium-magnesium salt of phytic acidxe2x80x94the hexaphosphate ester of inositol.
Phosphorus Deposition: The percent of phosphorus digested by an animal which is absorbed and is deposited in the body of the animal (bones, scales, flesh, etc.) in the growth processes. The undigested fraction of the P in the diet is excreted in the feces. xe2x80x9cPhosphorus Depositionxe2x80x9d has the same meaning as xe2x80x9cPhosphorus Retentionxe2x80x9d.
Protein Efficiency Ratio: As used herein, the protein efficiency ratio in a feeding study is the wet weight gain divided by the protein intake. The diets that utilized the protein in the diet the best would have the highest number.
Specific Growth Rate: As used herein, the specific growth rate is determined by dividing the weight gained by the number of days. Diets exhibiting the highest calculated number would have the best growth rate.
White Protein Gluten Meal: A protein feed ingredient derived by the wet milling of white corn hybrids in which the grain is typically separated into five separate segments consisting of protein, starch, fiber, embryo, and steep water. The white protein gluten meal is derived mainly from the endosperm fraction of the kernel and has protein levels higher than 70% (on a dry substance basis). This protein fraction is highly digestible, has low phosphorus with a range of about 0.30% to about 0.60%, and contains very low beta-carotene levels with a range of between about 0.03 mg/Kg to about 3.0 mg/Kg and/or low xanthophyll levels with a range of between about 2.5 mg/Kg to about 80 mg/Kg.
Xanthophyll content: The total amount of yellow carotenoid pigment.
The development of the white protein gluten meal is the result of identifying a specific white hybrid called 1851W that not only has special intrinsic grain traits, but also generates value-added co-products through industrial wet milling.
The corn hybrid, 1851W, is a proprietary single cross, white corn developed by Wilson Genetics, L.L.C. The hybrid was selected from various inbred crosses based upon one or more grain characteristics, e.g., color, yield, kernel hardness, cap smoothness, kernel size and shape uniformity, the protein characteristics of the endosperm, and the character of the starch. The intrinsic grain traits in 1851W includes higher grain protein levels ranges from about 9.3% to about 11.7%, dense starch that has a unique combination of 50.2% amylose and 49.8% amylopectin, increased endosperm protein (9.05%) that is very digestible, low endosperm phosphorus levels (0.07%), and high total dietary fiber (10.7%) on a dry substance basis. These traits can be selected in other white corn hybrids using conventional breeding techniques.
Contrary to the expectation that white corn performs poorly, if at all, in wet milling applications, the 1851W hybrid unexpectedly exhibited superior performance with standard wet milling methods. More specifically, the grain components of 1851W (starch, fiber, protein, and embryo) separated cleaner and easier than other white corn, and as well as many varieties of yellow dent corn, resulting in purer extracted products. It is believed that the superior performance of the 1851W hybrid with standard wet milling methods is due to the special grain traits of the 1851W hybrid. Other hybrids tested having these same special grain traits also have had superior performance with standard wet milling methods.
The deposition of minerals within the corn kernel is important when considering the separation of the kernel into components during the wet mill processing. In the 1851W grain, the endosperm contains 0.07% phosphorus while the embryo contains 0.79%. Consequently, if the embryo can be extracted intact and separated from the endosperm, then gluten, which is basically protein, derived from the endosperm, has a low phosphorus content.
The unexpected combination of the grain characteristics of 1851W, and with its wet milling attributes, has generated white protein products that have novel use applications in the feed industry.